EFFECTS OF SOME ORGANIC SOLVENTS ON GP lb AND ACTIN-BINDING PROTEIN IN BLOOD PLATELETS

1987 ◽  
Author(s):  
A M Aakhus ◽  
N O Solum ◽  
I Hagen
Keyword(s):  
Organic Solvents ◽  
Von Willebrand Factor ◽  
Early Stage ◽  
Binding Protein ◽  
Blood Platelets ◽  
Cytoskeletal Proteins ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Von Willebrand ◽  
Willebrand Factor

Effects on filamentous proteins appear to be a central phenomenon in the neuronal toxic effects of organic solvents. We have therefore compared the effects of some organic solvents (particularly isopropylalcohol, IPA) to the previously observed effects of dibucaine (DBC) on platelet cytoskeletal proteins. Incubation of platelets with 6% IPA at 37° C, like DBC, initiates a degradation of actin-binding protein (ABP) as substrate for a calcium activated protease (CAP), shown by SDS-PAGE. IPA leads to an increase followed by a decrease in bovine von Willebrand factor-induced agglutination. The decrease is accompanied by a release of glycocali-cin from the GP lb α-chain. The process was also studied using CIE of Triton X-100 extracts of platelets against antiserum to glycocalicin. Incubation of platelets with IPA before extraction in the presence of 4.2 mM leupeptin leads to a time-dependent transformation of GP Ib-related immunoprecipitates from that of the slow-migrating peak III complex (probably between ABP and GP lb) to the faster migrating GP Ib-precipitate. Our working hypothesis is that IPA induces an activation of the CAP by mobilizing calcium. This leads to degradation of ABP and liberation of GP lb from the cytoskeleton accompanied by an increased tendency for agglutination. The following decrease is explained by degradation of the glycocalicin part of the GP lb enchain which contains the binding-site for von Willebrand factor. We conclude that IPA has a similar effect on GP lb and ABP as DBC. Preliminary studies with 1% DMSO and 0,005% toluene at 37° C revealed that these organic solvents have some similar effects on platelets as described for IPA. Possibly the described effects are characteristic of certain cells at an early stage in a process ultimately leading to cell lysis.

Cytoskeletal regulation of the platelet glycoprotein Ib/V/IX–von Willebrand factor interaction

Blood ◽  
2000 ◽  
Vol 96 (10) ◽  
pp. 3480-3489 ◽  
Author(s):  
Nayna Mistry ◽  
Susan L. Cranmer ◽  
Yuping Yuan ◽  
Pierre Mangin ◽  
Sacha M. Dopheide ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Von Willebrand Factor ◽  
Cho Cells ◽  
Actin Polymerization ◽  
Cytochalasin D ◽  
Actin Binding ◽  
Platelet Glycoprotein ◽  
Actin Binding Protein ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Shear-induced binding of von Willebrand factor (vWf) to the platelet glycoprotein (GP) Ib/V/IX complex plays a key role in initiating platelet adhesion and aggregation at sites of vascular injury. This study demonstrated that pretreating human platelets with inhibitors of actin polymerization, cytochalasin D or latrunculin B, dramatically enhances platelet aggregation induced by vWf. The effects of these inhibitors were specific to the vWf-GPIbα interaction because they enhanced vWf-induced aggregation of Glanzmann thrombasthenic platelets and Chinese hamster ovary (CHO) cells transfected with GPIb/V/IX. Moreover, cytochalasin D enhanced the extent of platelet aggregation induced by high shear stress (5000 s−1) and also lowered the shear threshold required to induce aggregation from 3000 s−1 to as low as 500 s−1. Studies of CHO cells expressing GPIbα cytoplasmic tail truncation mutants that failed to bind actin-binding protein-280 (deletion of residues 569-610 or 535-568) demonstrated that the linkage between GPIb and actin-binding protein-280 was not required for vWf-induced actin polymerization, but was critical for the enhancing effects of cytochalasin D on vWf-induced cell aggregation. Taken together, these studies suggest a fundamentally important role for the cytoskeleton in regulating the adhesive function of GPIb/V/IX.

Cytoskeletal regulation of the platelet glycoprotein Ib/V/IX–von Willebrand factor interaction

Blood ◽  
2000 ◽  
Vol 96 (10) ◽  
pp. 3480-3489 ◽  
Author(s):  
Nayna Mistry ◽  
Susan L. Cranmer ◽  
Yuping Yuan ◽  
Pierre Mangin ◽  
Sacha M. Dopheide ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Von Willebrand Factor ◽  
Cho Cells ◽  
Actin Polymerization ◽  
Cytochalasin D ◽  
Actin Binding ◽  
Platelet Glycoprotein ◽  
Actin Binding Protein ◽  
Von Willebrand ◽  
Willebrand Factor

Shear-induced binding of von Willebrand factor (vWf) to the platelet glycoprotein (GP) Ib/V/IX complex plays a key role in initiating platelet adhesion and aggregation at sites of vascular injury. This study demonstrated that pretreating human platelets with inhibitors of actin polymerization, cytochalasin D or latrunculin B, dramatically enhances platelet aggregation induced by vWf. The effects of these inhibitors were specific to the vWf-GPIbα interaction because they enhanced vWf-induced aggregation of Glanzmann thrombasthenic platelets and Chinese hamster ovary (CHO) cells transfected with GPIb/V/IX. Moreover, cytochalasin D enhanced the extent of platelet aggregation induced by high shear stress (5000 s−1) and also lowered the shear threshold required to induce aggregation from 3000 s−1 to as low as 500 s−1. Studies of CHO cells expressing GPIbα cytoplasmic tail truncation mutants that failed to bind actin-binding protein-280 (deletion of residues 569-610 or 535-568) demonstrated that the linkage between GPIb and actin-binding protein-280 was not required for vWf-induced actin polymerization, but was critical for the enhancing effects of cytochalasin D on vWf-induced cell aggregation. Taken together, these studies suggest a fundamentally important role for the cytoskeleton in regulating the adhesive function of GPIb/V/IX.

scholarly journals Disappearance of actin binding protein from human blood platelets during storage

FEBS Letters ◽  
1979 ◽  
Vol 102 (2) ◽  
pp. 257-260 ◽  
Author(s):  
Frank A. Robey ◽  
Catherine M. Freitag ◽  
G.A. Jamieson
Keyword(s):  
Human Blood ◽  
Binding Protein ◽  
Blood Platelets ◽  
Actin Binding ◽  
Actin Binding Protein ◽  
Human Blood Platelets

scholarly journals Identification of membrane proteins mediating the interaction of human platelets

1980 ◽  
Vol 86 (1) ◽  
pp. 77-86 ◽  
Author(s):  
D Phillips ◽  
L Jennings ◽  
H Edwards
Keyword(s):  
Actin Filaments ◽  
Binding Protein ◽  
Membrane Surface ◽  
Direct Interaction ◽  
Cytoskeletal Proteins ◽  
Actin Binding ◽  
Insoluble Residue ◽  
Membrane Glycoproteins ◽  
Actin Binding Protein ◽  
Activated Platelets

Membrane glycoproteins that mediate platelet-platelet interactions were investigated by identifying those associated with the cytoskeletal structures from aggregated platelets. The cytoskeletal structures from washed platelets, thrombin-activated platelets (platelets incubated with thrombin in the presence of mM EDTA to prevent aggregation) and thrombin- aggregated platelets (platelets activated in the presence of mM Ca(++) were prepared by first treating platelet suspensions with 1 percent Triton X-100 and 5 mM EGTA and then isolating the insoluble residue by centrifugation. The readily identifiable structures in electron micrographs of the residue from washed platelets had the shape and dimensions of actin filaments. Analysis of this residue from washed platelets had the shape and dimensions of actin filaments. Analysis of this residue by SDS gel electrophoresis showed that it consisted primarily of three proteins: actin (mol wt = 43,000), myosin (mol wt = 200,000) and a high molecular weight polypeptide (mol wt = 255,000) which had properties indentical to actin-binding protein (filamin). When platelets are activated with thrombin in the presence of EDTA to prevent aggregation, there was a marked increase in the amount of insoluble precipitate in the subsequent Triton extraction. Transmission electron microscopy showed that this residue not only contained the random array of actin filaments as seen above, but also organized structures from individual platelets which appeared as balls of electron-dense filamentous material approximately 1mum in diameter. SDS polyacrylamide gel analysis of the Triton residue of activated platelets showed that this preparation contained more actin, myosin and actin-binding protein than that from washed platelets plus polypeptides with mol wt of 56,000 and 90,000 and other minor polypeptides. Thus, thrombin activation appeared to increase polymerization of actin in association with other cytoskeletal proteins into structures that are observable after Triton extraction. The cytoskeletal structures from thrombin-aggregated platelets were similar to those from thrombin-activated platelets, except that the structural elements from individual platelets remained aggregated rather than randomly dispersed in the actin filaments. This suggested that the membrane components that mediate the direct interaction of platelets were in Triton residue from aggregated platelets. Only a small percentage of the membrane surface proteins and glycoproteins were found in the cytoskeletal structures from either washed platelets or thrombin-activated platelets. In contrast, the aggregated cytoskeletal structures from thrombin-aggregated platelets contained membrane glycoproteins IIb (26 percent of the total in pre-extracted platelets) and III (14 percent), suggesting that one or both of these glycoproteins participate in the direct interaction of platelets during aggregation.

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